Eduardo Cesar Tondo

Identification of multiple strains of Staphylococcus aureus colonizing nasal mucosa of food handlers

Abstract The occurrence of multiple strains of Staphylococcus aureus colonizing the nasal mucosa of food handlers was investigated. Samples were collected from the anterior nares of 47 food handlers and were streaked on plates of Baird-Parker agar. From each plate, three typical colonies of S. aureus , with similar morphologies, were picked up and identified by biochemical tests. S. aureus isolates were typed by antibiotic susceptibility and pulsed-field gel electrophoresis (PFGE). Results indicated nasal carriage of S. aureus in 14 (30%) of the personnel. The isolated micro-organisms were resistant to penicillin and amoxicillin+clavulanic acid (70% and 45% of the isolates, respectively). All isolates were sensitive to vancomycin, rifampicin, cephalothin, oxacillin, chloramphenicol, gentamycin, and ofloxacin. Typing methods indicated that 11 among the 14 food handlers harbored multiple strains of S. aureus per carrier. Such results demonstrate that multiple isolates of S. aureus need to be strain-typed per food handler when attempts are made to identify sources of food poisoning, in epidemiological studies and in investigations of food contamination sources.

Serological Characterization and Prevalence of spvR Genes in Salmonella Isolated from Foods Involved in Outbreaks in Brazil

Salmonella strains (n = 75) isolated from foods involved in foodborne outbreaks occurred in Rio Grande do Sul State, Brazil, during 1999 and 2000 were studied. Strains were serotyped and submitted to PCR analysis to verify the prevalence of Salmonella plasmid virulence (spvR) regulatory gene. Among the 75 isolates, 73 (97%) were classified as Salmonella enterica serovar Enteritidis. All of the Salmonella strains isolated in 1999 were classified as serotype Enteritidis, whereas in 2000 two isolates were serotyped as Salmonella Derby and Salmonella Typhimurium. Regarding the prevalence of spvR gene, 62 strains (82.7%) were PCR positive, and a positive correlation (P < 0.05) between the strains of Salmonella Enteritidis and the presence of spvR gene was demonstrated, which suggests that this gene is a characteristic of the Salmonella Enteritidis analyzed.

Salmonellosis in Rio Grande do Sul, Brazil, 1997 to 1999

Salmonella spp. was the major cause of reported foodborne diseases in the last years in the State of Rio Grande do Sul, South of Brazil. Epidemiological data on salmonellosis, occurred in the period of 1997 to 1999 and supplied by the Division of Sanitary Surveillance (DVS/RS), were analysed according to the following factors: total number of confirmed outbreaks, number of people involved, outbreaks incidence according to the period of the year, age and sex of involved people, food vehicle, food storage conditions, local where the disease occurred, and possible causes of the outbreaks. The results indicated that 8217 people were involved, and 1557 had to be hospitalized. The highest number of outbreaks occurred during springtime and the principal age group affected was between 16 and 50 years. The most common food vehicle was salad prepared with homemade mayonnaise (42.45%). The principal causes of salmonellosis were raw-materials not submitted to regulatory inspection (22.92%), mainly eggs, and foods maintained at room temperature for more than 2 hours (20.55%). The majority of the outbreaks occurred in private homes (43.70%) and commercial food establishments (25.21%).

Microbiological quality and safety assessment of lettuce production in Brazil

The microbiological quality and safety of lettuce during primary production in Brazil were determined by enumeration of hygiene indicators Escherichia coli, coliforms and enterococci and detection of enteric pathogens Salmonella and E. coli O157:H7 in organic fertilizers, soil, irrigation water, lettuce crops, harvest boxes and workers hands taken from six different lettuce farms throughout the crop growth cycle. Generic E. coli was a suitable indicator for the presence of Salmonella and E. coli O157:H7, while coliforms and enterococci were not. Few pathogens were detected: 5 salmonellae and 2 E. coli O157:H7 from 260 samples, of which only one was lettuce and the others were manure, soil and water. Most (5/7) pathogens were isolated from the same farm and all were from organic production. Statistical analysis revealed the following environmental and agro-technical risk factors for increased microbial load and pathogen prevalence in lettuce production: high temperature, flooding of lettuce fields, application of contaminated organic fertilizer, irrigation with water of inferior quality and large distances between the field and toilets. Control of the composting process of organic fertilizers and the irrigation water quality appear most crucial to improve and/or maintain the microbiological quality and safety during the primary production of lettuce.

Identification of heat stable protease of Klebsiella oxytoca isolated from raw milk

Aims:  The aim of this study was to identify and characterize heat stable proteinases of psychrotrophic proteolytic bacteria isolated from raw milk.

Evaluation of an indirect ELISA for the detection of Salmonella in chicken meat

In this work, an indirect ELISA based on a monoclonal antibody (MAb) specific for an outer membrane protein of Salmonella enterica serovar Enteritidis was used for detection of Salmonella in 154 samples of chicken meat. Its efficiency was determined through comparison with the results obtained from the conventional method. The prevalence of samples contaminated with Salmonella was 23% with the conventional culture method, and 26% with the ELISA. From thirty-five samples positive for Salmonella by the conventional method, 33 were also positive by ELISA. Seven other samples were only positive in the ELISA. Comparison of the results obtained in the two methods showed an ELISA sensitivity and specificity of 94%, and positive and negative predictive values of 82% and 98% respectively. The serotyping of the isolates revealed 31 Salmonella enterica serovar Enteritidis, 2 Salmonella enterica serovar Heidelberg, 1 Salmonella enterica serovar Choleraesuis and 1 Salmonella enterica sorovar 6,7:-:-.

Assessing the epidemiological data of Staphylococcus aureus food poisoning occurred in the State of Rio Grande do Sul, Southern Brazil

Staphylococcal food poisoning is one of the most frequent foodborne illnesses worldwide and it is caused by the ingestion of food contaminated with enterotoxins produced by some strains of Staphylococcus (S.) aureus. In the State of Rio Grande do Sul (RS), Southern Brazil, S. aureus has been identified as the second most frequent agent of foodborne illnesses in the last two decades. The aim of the present study was to assess and analyse the epidemiological data of S. aureus food poisoning occurred in the State of RS during the years of 2000 to 2002. The official records of epidemiological investigations carried out by the Sanitary Surveillance Services of the State of RS were analysed. Among foodborne outbreaks for which aetiology was determined, S. aureus was identified as the responsible agent of 57 foodborne outbreaks, being 42 (74%) confirmed by microbiological analyses and 15 (26%) confirmed by clinical symptoms and/or epidemiological data. Staphylococcal outbreaks were responsible for the exposition of 5,991 persons, of which 1,940 (32%) were interviewed by the Sanitary Surveillance officers. The most affected age group corresponded to people with 20 to 49 years old (48%), where men (48%) and women (52%) were affected similarly. The main involved food vehicles were meats servings (35%), followed by pastries (25%), cheese (23%), pasta (11%) and potato salad with homemade mayonnaise (11%). The majority of the outbreaks occurred inside private homes (33%) followed by commercial food establishments (28%). Inadequate control of temperature and failures in general hygiene practices were identified as the main factors responsible for the outbreaks. In conclusion, S. aureus was an important food poisoning etiological agent in the State of RS during 2000 to 2002 and its prevention depends on control measures involving different parts of the food chain.

Risk Factors for Salmonella, Shiga Toxin-Producing Escherichia coli and Campylobacter Occurrence in Primary Production of Leafy Greens and Strawberries.

The microbiological sanitary quality and safety of leafy greens and strawberries were assessed in the primary production in Belgium, Brazil, Egypt, Norway and Spain by enumeration of Escherichia coli and detection of Salmonella, Shiga toxin-producing E. coli (STEC) and Campylobacter. Water samples were more prone to containing pathogens (54 positives out of 950 analyses) than soil (16/1186) and produce on the field (18/977 for leafy greens and 5/402 for strawberries). The prevalence of pathogens also varied markedly according to the sampling region. Flooding of fields increased the risk considerably, with odds ratio (OR) 10.9 for Salmonella and 7.0 for STEC. A significant association between elevated numbers of generic E. coli and detection of pathogens (OR of 2.3 for STEC and 2.7 for Salmonella) was established. Generic E. coli was found to be a suitable index organism for Salmonella and STEC, but to a lesser extent for Campylobacter. Guidelines on frequency of sampling and threshold values for E. coli in irrigation water may differ from region to region.

Adhesion and biocides inactivation of Salmonella on stainless steel and polyethylene.

The adhesion of Salmonella (S.) strains to stainless steel and polyethylene and their inactivation by biocides used in food industry was investigated. Coupons of stainless steel and polyethylene were immersed in bacterial suspensions of S. Enteritidis, S. Typhimurium, and S. Bredeney during 15, 30, and 60 minutes, and submitted to different concentrations of peracetic acid (PAA), sodium hypochlorite (NaOCl), and quaternary ammonium (Quat) sanitizers. Hydrophobicity of the surfaces was evaluated by contact angle measurements using the sessile drop method and bacterial adhesion was accompanied through bacterial counts and scanning electron microscopy (SEM). Results indicated that the three serovars of Salmonella presented similar adhesion to both materials (5.0 to 6.5 log cfu cm -2 ). The time of exposure did not influence the counts of adhered cells on both surfaces, however SEM revealed larger clusters of S. Enteritidis on both materials, not found for the other serovars. S. Enteritidis presented lower sessile drop angle on polyethylene, indicating hydrophilic properties of this material. The biocides were not able to inactivate all the microorganisms adhered on both surfaces. At least 1 log cfu cm -2 of all serovars tested remained viable after the exposure to different biocide concentrations. In general, higher counts of survivors were observed on polyethylene disinfected with different concentrations of biocides. S. Bredeney e S. Typhimurium were more resistant than S. Enteritidis to PAA, whilst S. Enteritidis presented smaller reduction rates to NaOCl. This last biocide was able to reduce Salmonella counts in approximately 3.0 to 4.0 log cm -2 . When adhered to polyethylene, the serovars S. Typhimurium and S. Enteritidis were more resistant to Quat than S. Bredeney in all concentrations tested, and the numbers of S. Enteritidis remained almost unaltered. On stainless steel disinfected by Quat, S. Bredeney presented higher numbers of survivors.

ANTIMICROBIAL RESISTANCE AND PCR-RIBOTYPING OF SHIGELLA RESPONSIBLE FOR FOODBORNE OUTBREAKS OCCURRED IN SOUTHERN BRAZIL

Little information about Shigella responsible for foodborne shigellosis is available in Brazil. The present study aimed to investigate the antimicrobial resistance and PCR-ribotyping patterns of Shigella isolates responsible for foodborne outbreaks occurred in Rio Grande do Sul State (RS), Southern Brazil in the period between 2003 and 2007. Shigella strains (n=152) were isolated from foods and fecal samples of victims of shigellosis outbreaks investigated by the Surveillance Service. Identification of the strains at specie level indicated that 71.1% of them were S. flexneri, 21.5% S. sonnei, and 0.7% S. dysenteriae. Ten strains (6.7%) were identified only as Shigella spp. An increasing occurrence of S. sonnei was observed after 2004. Most of the strains were resistant to streptomycin (88.6%), followed by ampicillin (84.6%), and sulfamethoxazole/trimethoprim (80.5 %). Resistant strains belonged to 73 patterns, and pattern A (resistance to ampicillin, sulfamethoxazole/trimethoprim, tetracycline, streptomycin, chloramphenicol, and intermediate resistance to kanamycin) grouped the largest number of isolates (n=36). PCR-ribotyping identified three banding patterns (SH1, SH2, and SH3). SH1 grouped all S. flexneri and SH2 grouped all S. sonnei. The S. dysenteriae strain belonged to group SH3. According to the results, several Shigella isolates shared the same PCR-rybotyping banding pattern and the same resistance profile, suggesting that closely related strains were responsible for the outbreaks. However, other molecular typing methods need to be applied to confirm the clonal relationship of these isolates.